A Multimethod Analysis to Assess Locomotor Capabilities in Stem Tetrapods from Blue Beach (Tournaisian; Early Carboniferous), Nova Scotia

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2020-01 A multimethod analysis to assess locomotor capabilities in stem tetrapods from Blue Beach (Tournaisian; Early Carboniferous), Nova Scotia

Lennie, Kendra Ilana

Lennie, K. I. (2020). A multimethod analysis to assess locomotor capabilities in stem tetrapods from Blue Beach (Tournaisian; Early Carboniferous), Nova Scotia (Unpublished master's thesis). University of Calgary, Calgary, AB. http://hdl.handle.net/1880/111587 master thesis

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A multimethod analysis to assess locomotor capabilities in stem tetrapods from Blue Beach

(Tournaisian; Early Carboniferous), Nova Scotia

Kendra Ilana Lennie

A THESIS

SUBMITTED TO THE FACULTY OF GRADUATE STUDIES

IN PARTIAL FULFILMENT OF THE REQUIREMENTS FOR THE

DEGREE OF MASTER OF SCIENCE

GRADUATE PROGRAM IN BIOLOGICAL SCIENCES

CALGARY, ALBERTA

JANUARY, 2020

© Kendra Ilana Lennie 2020 Abstract In vertebrate evolution the fin-to-limb transition was an important precursor to the diversification and radiation of terrestrial animals into novel environments. This transition began in the Devonian and continued through the Carboniferous and involved physiological and biomechanical changes. I used a multi-method approach to assess external and internal limb bone features to evaluate Early Carboniferous (Tournaisian) limb bones from Blue Beach and associated them with aquatic to terrestrial lifestyles. Tournaisian tetrapod material was collected at Blue Beach located near Hantsport, Nova Scotia, but much of it has not been formally described because the disarticulated and isolated tetrapod elements made identification to the species level difficult. In this thesis I described new morphotypes attributable to the family level which are used in the following chapters. Once the external morphology of the Blue Beach bones was described I compared them with the femora of extant aquatic, amphibious, and terrestrial tetrapods to evaluate which locomotor behaviour the fossil femora most resembled. I additionally examined cross-sectional bone profiles of Blue Beach tetrapod femora to infer lifestyle. Midshaft analyses relied on a single two-dimensional image to represent a dynamically structured bone so

I also used a novel method for assessing three-dimensional trabecular data to qualitatively and quantitatively infer lifestyle from the Blue Beach femora. From the various analyses of internal and external bone morphology it was clear that external bone features of modern and early fossil tetrapod femora are dissimilar, which lead to difficulties in drawing conclusions based off external qualitative data. Internal data, from two-dimensional midshaft and three-dimensional trabecular structures, produced quantitative results that lead to the same conclusion, that the Blue

Beach femora are consistent with those of aquatic animals. This implies that the initial diversification of the tetrapod body plans present in the Early Carboniferous was not the result of terrestrialization but appears to have preceded it.

Preface This thesis is original, unpublished, work by the author, K. Lennie.

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Acknowledgments I would like to thank my supervisor Jason Anderson for helping me revise countless drafts and being patient with me for the last two and a half years. I would also like to thank my committee members Jessica Theodor, Sarah Manske, and Heather Jamniczky, whose support and suggestions (especially about trabecular bone analysis) was critical to the success of my project.

I would like to thank NSERC for funding a Discovery Grant to Jason Anderson, which made this work possible. Additionally I’d like to thank Natalie Reznikov for training on the Bone Analysis module of Dragonfly ORS (Montreal), Katherine Ogdon and Tim Fedak for access to the specimens at the Nova Scotia Museum, Chris Mansky and Sonja Wood for access and preparation work on the specimens from the Blue Beach Fossil Museum, and Johanne Kerr for help finding and accessing elements from Eusthenopteron at Musée d’Histoire Naturelle de

Miguasha. I want to thank my lab mates Jason Pardo, Conrad Wilson, James Campbell, Ramon

Nagesan, Amber Whitbone, and Jamey Creighton, and Keith Johnstone for many informative and thought-provoking discussions, for listening to my constant complaining about muscles, and for providing feedback on many written applications and assignments. I would finally especially like to thank my parents for pushing me to do my best and supporting me in all my endeavours.

Dedication I dedicate this thesis to all the girls and women who are defying traditional gender roles in science and sport. Never let anyone tell you should not or cannot do something because you are a girl.

Table of Contents Abstract ...... ii Preface ...... iii Acknowledgments...... iv Dedication ...... v Table of Contents ...... vi List of Tables ...... ix List of Figures ...... x Table of extant animals ...... xiv Abbreviations ...... xv Institutional ...... xv Anatomical ...... xv Variables ...... xv Chapter 1: Introduction ...... 1 Introduction ...... 1 Historic hypotheses ...... 2 Romer’s Gap ...... 2 Blue Beach ...... 3 Historical proxies for locomotion ...... 6 Predictions ...... 11 Literature cited ...... 13 Chapter 2: Descriptions of new tetrapod limb elements from Blue Beach, Nova Scotia ...... 23 Introduction ...... 23 Abbreviations ...... 23 Geological context ...... 24 Material and Methods ...... 25 Results ...... 25 Humeri ...... 25 Ulna ...... 26 Radii ...... 27 Femora ...... 29 Fibulae ...... 41 Tibiae ...... 44 Discussion ...... 48

Conclusion ...... 50 Literature Cited ...... 51 Chapter 3: Musculature comparisons using extant taxa ...... 56 Introduction ...... 56 Materials and Methods ...... 59 Results ...... 62 Large Osteological Features ...... 63 Discussion ...... 68 Gross patterns in bone morphology in representative extant taxa and comparison to the Blue Beach material...... 68 Extensive pitting of early tetrapod femora ...... 72 Conclusions ...... 73 Literature Cited ...... 75 Chapter 4: Midshaft profiles ...... 82 Introduction ...... 82 Abbreviations ...... 86 Materials and Methods ...... 87 Results ...... 93 NSM.005.GF.045.044 ...... 94 NSM.005.GF.045.047-Distal ...... 94 NSM.007.GF.004.630 ...... 95 NSM.005.GF.045.048-A ...... 96 RM.20.6711 ...... 96 YPM.PU.20103 ...... 96 Discussion ...... 98 Conclusion ...... 100 Literature Cited ...... 101 Chapter 5: Trabecular Analysis ...... 109 Introduction ...... 109 Wolff’s law ...... 112 Anisotropy ...... 112 Fin-to-limb ...... 114 Abbreviations ...... 115 Locality ...... 116 Materials and Methods ...... 116

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Results ...... 126 Extant terrestrial taxa ...... 126 Extant aquatic taxa ...... 130 Fossil bones ...... 134 Principal Component Analysis ...... 143 Discussion ...... 147 Femora ...... 147 Forelimb elements ...... 152 General observations ...... 153 Principal Component Analysis ...... 153 Conclusions ...... 156 Literature cited ...... 157 Chapter 6: Summary and Conclusions...... 170 Chapter 1 summary ...... 170 Chapter 2 summary ...... 170 Chapter 3 summary ...... 171 Chapter 4 summary ...... 172 Chapter 5 summary ...... 174 Concluding Remarks ...... 176 Literature Cited ...... 177 Literature Cited ...... 180 Appendix A ...... 195 Appendix B ...... 197 Appendix C ...... 229 Appendix D ...... 242 Appendix E ...... 266 Appendix F ...... 283 Appendix G ...... 288

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List of Tables

Table 2.1. Borrowed Blue Beach elements and their attributed taxonomic morphologies and lifestyles. . 47 Table 4.1 Femora used for midshaft profiles, and their historical locomotor behaviour associations...... 89 Table 4.2. ImageJ threshold functions and values of midshaft slices...... 91 Table 4.3. Angular Compactness Profiles of Blue Beach Femora Midshafts ...... 95 Table 5.1. Variables of whole bone analysis ...... 124 Table 5.2. Summary of Primary Component Analyses ...... 144 Table 5.3. PCA loadings ...... 145 Table S.0.1 PCA summary statistics ...... 283 Table S.0.2 PCA loadings ...... 283 Table S.1 PCA no Felis summary ...... 288 Table S.2 PCA no Felis loadings ...... 288

List of Figures Figure 1.1. The outgroups and sister group to early tetrapods that are required to infer soft tissues using the extant phylogenetic bracket...... 10 Figure 2.1. Micro computed tomography (µCT) images of NSM.007.GF.004.733: right ulna. Tulerpeton- like ulna in (A) extensor, (B) anterior, (C) flexor, (D) posterior, (E) proximal and (F) distal view. mr, mesial ridge. Scalebar 10mm...... 27 Figure 2.2. µCT images of NSM.014.GF.036.02, right Acanthostega-like radius (A-F) and NSM.007.GF.004.767, left radius (G-L) in (A, G) extensor, (B, H) posterior, (C, I) flexor, (E, K) proximal and (F, L) distal view. vrc, ventral radial crest. Scalebar 10mm...... 28 Figure 2.3. µCT images of NSM.005.GF.045.048-A, left embolomerous femur (A-F), and RM.20.5222, a right Acanthostega-like) femur (G-L) in (A, G) extensor, (B, H) anterior, (C, I) flexor, (D, J) posterior, (E, K) proximal, and (F, L) distal views. ab, adductor blade; ac, adductor crest; icf, intercondylar fossa; itf, intertrochanteric fossa; pa, popliteal area. Scalebar 10mm...... 30 Figure 2.4. µCT images of NSM.007.GF.004.630 a right Tulerpeton-like femur in (A) extensor, (B) anterior, (C) flexor, (D) posterior, (E) proximal, and (F) distal view. ab, adductor blade; ac, adductor crest; cp, central prominence; ff, fibular facet; fo, fibular fossa; itf, intertrochanteric fossa; pa, popliteal area; tf, tibial facet. Scalebar 10mm...... 32 Figure 2.5. µCT images of YPM.PU.20103 a right Tulerpeton-like/whatcheeriid femur (A-F) and NSM.005.GF.045.042 a left Tulerpeton-like femur (G-L) in (A,G) extensor, (B,H) anterior, (C,I) flexor, (D,J) posterior, (E,K) proximal, and (F,L) distal views. scalebar 10mm...... 34 Figure 2.6. µCT images (meshes) of YPM.PU.20103 (A-D) and NSM.005.GF.045.042 (E-H) in (A,E) extensor, (B,F) anterior, (C,G) flexor and, (D,H) posterior views. ab, adductor blade; ac, adductor crest; der, dorsal elongate ridge; ff, fibular facet; fo, fibular fossa; icf, intercondylar fossa; it, internal trochanter; itf, intertrochanteric fossa; pa, popliteal area; tf, tibial facet. Scalebar 10mm...... 36 Figure 2.7. µCT images of NSM.005.GF.045.044, distal end of a left Ossinodus-like femur (A-E), and NSM.005.GF.045.047 proximal (K-O) and distal (F-J) segments in (A,F,K) extensor, (B,G,L) anterior, (C,H,M) flexor, (D,I,N) posterior, (E,J) distal and (O) proximal views. ab, adductor blade; ac, adductor crest; cp, central prominence; ff, fibular facet; fo, fibular fossa; ft, fourth trochanter; itf, intertrochanteric fossa; pa, popliteal area; tf, tibial facet. Scalebar 10mm...... 39 Figure 2.8. µCT images of NSM.005.GF.045.061 left Pederpes-like fibula (A-F) in and left fibula NSM.005.GF.045.063 (G-L) in (A,G) extensor, (B,H) posterior, (C,I) flexor, (D,J) anterior, (E,K) proximal, and (F,L) distal views. Scalebar 10mm...... 43 Figure 2.9. µCT images of left tibiae RM.20.2555 (A-F) and NSM.005.GF.045.320 (G-L) in (A,G) extensor, (B,H) anterior, (C,I) flexor, (D,J) posterior, (E,K) proximal and (F,L) distal views. Scalebar 10mm ...... 45 Figure 3.1 Cladogram showing nested groups more closely related to each other than the taxa to the left. For example, birds and crocodiles form a sister group to lepidosaurs, and are more closely related to each other than they are to lepidosaurs. Lepidosaurs, crocodiles, and birds form a sister groups with mammals, and are more closely related to each other than they are to mammals. And so on. The silhouettes illustrate there is a great deal of variation in body plans, and all major extant groups have some taxa that have undergone a secondary transition to an aquatic or amphibious lifestyle...... 57 Figure 3.2. Progression of sampled aquatic to amphibious to terrestrial femora sampled. µCT images ofA, femur of Necturus (aquatic). B, femur of Cynops (amphibious). C, femur of Uromastyx (terrestrial). D, femur of Eublepharis (terrestrial). Scale bars represent 1mm...... 63

Figure 3.3. Comparison of topological similarities between the femora of Cynops (a,c) and Blue Beach YPM.PU.20103 (b,d). Yellow appears to resemble an intertrochanteric fossa, blue the popliteal area, and purple a fibular fossa. Red represents well developed ridges ...... 66 Figure 3.4. Tetrapod femur NSM.007.GF.004.630. A, green arrows indicate where large rugose pits extend through the cortical bone. B, same pits in A visible in transaxial sections crossing through the cortical layer. C, same pits viewed in longitudinal sections crossing the cortical layer...... 69 Figure 3.5. Femur NSM.007.GF.004.630. Green arrows are pointing to small “pits” found across blue beach taxa, similar in size to those described by Bishop (2014). B, same pits in A visible in transaxial sections crossing through the cortical layer. C, same pits viewed in longitudinal sections crossing the cortical layer...... 71 Figure 4.1 Bone Profiler analysis. A, Schematic of the 51 ring and 6-degree increments analysed by bone profiler. B, Graphical representation of the four compactness parameters. Modified from Quemeneur et al. (2013)...... 88 Figure 4.2. Example of how a midshaft (RM.20.6711) was analysed in bone profiler. A, original midshaft image. B, midshaft after contrast was adjusted. C, binary midshaft after threshold applied. D, binary image uploaded into Bone Profiler and bone parameters automatically generated. E, global and angular output from Bone Profiler...... 93 Figure 4.3. Bone Profiler graphs illustrating the compactness profiles of the Blue Beach fossil Femora, with a platypus (MVZ:32885) included for comparison representing an aquatic bone profile, and cat representing terrestrial...... 94 Figure 4.4. Unmodified µCT (A,C,E,G, I, and K) and thresholded binary images (B,D,F,H, J, and L) of the midshafts of fossil femora...... 97 Figure 5.1. Coronal slice of the proximal femur of a juvenile cat. Highlighted in red is highly aligned trabecular structure that is indicative of the force transmitted from pelvis to femur in response to walking in a terrestrial environment (i.e., under gravitational forces)...... 111 Figure 5.2. Transaxial slice of fossil tibia NSM.014.GF.036.004. trabecular structure is highlighted in pink. A shows that trabecular structure is left with aligned edges where it is cut from the trabecular structure in segmentation. B shows how this effects anisotropy calculations at the edges where these cuts get analysed and produce areas of high anisotropy. C shows the inverted trabecular area (grey-blue) that will be overwritten on the anisotropy heat map. D shows the more accurate overwritten anisotropy heat map...... 123 Figure 5.3. Anisotropy vector maps and histograms of extant terrestrial humeri. A-D Eublepharis humerus. B, vector map of all anisotropy. C, Vector map of anisotropy 0.65 and above. D, Vector map of the middle region in the coronal plane with anisotropy 0.65 and above. E, Anisotropy frequency histogram of Eublepharis humerus. F-I, Uromastyx humerus. G, vector map of all anisotropy. H, Vector map of anisotropy 0.65 and above. I, Vector map of the middle region in the coronal plane with anisotropy 0.65 and above. J, Anisotropy frequency histogram of Uromastyx humerusFigure 5.2. Transaxial slice of fossil tibia NSM.014.GF.036.004. trabecular structure is highlighted in pink. A shows that trabecular structure is left with aligned edges where it is cut from the trabecular structure in segmentation. B shows how this effects anisotropy calculations at the edges where these cuts get analysed and produce areas of high anisotropy. C shows the inverted trabecular area (grey-blue) that will be overwritten on the anisotropy heat map. D shows the more accurate overwritten anisotropy heat map. . 123 Figure 5.3. Anisotropy vector maps and histograms of extant terrestrial humeri. A-D Eublepharis humerus. B, vector map of all anisotropy. C, Vector map of anisotropy 0.65 and above. D, Vector map of the middle region in the coronal plane with anisotropy 0.65 and above. E, Anisotropy frequency histogram of Eublepharis humerus. F-I, Uromastyx humerus. G, vector map of all anisotropy. H, Vector

xi map of anisotropy 0.65 and above. I, Vector map of the middle region in the coronal plane with anisotropy 0.65 and above. J, Anisotropy frequency histogram of Uromastyx humerus...... 127 Figure 5.4. Anisotropy vector maps and histograms of extant terretrial femora. A-D Eublepharis femur. B, vector map of all anisotropy. C, Vector map of anisotropy 0.65 and above. D, Vector map of the middle region in the coronal plane with anisotropy 0.65 and above. E, Anisotropy frequency histogram of Eublepharis femur. F-I, Uromastyx femur. G, vector map of all anisotropy. H, Vector map of anisotropy 0.65 and above. I, Vector map of the middle region in the coronal plane with anisotropy 0.65 and above. J, Anisotropy frequency histogram of Uromastyx femur. K-N, Felis femur. L, vector map of all anisotropy. M, vector map of anisotropy of 0.65 and above. N, vector map of the mid region of the bone in the coronal plane with anisotropy 0.65 or higher. O, Anisotropy frequency histogram of Felis femur...... 129 Figure 5.5. Anisotropy vector maps and histograms of extant aquatic humeri. A-D Cryptobranchus humerus. B, vector map of all anisotropy. C, Vector map of anisotropy 0.65 and above. D, Vector map of the middle region in the coronal plane with anisotropy 0.65 and above. E, Anisotropy frequency histogram of Cryptobranchus humerus. F-I, Ornithorhynchus humerus. G, vector map of all anisotropy. H, Vector map of anisotropy 0.65 and above. I, Vector map of the middle region in the coronal plane with anisotropy 0.65 and above. J, Anisotropy frequency histogram of Ornithorhynchus humerus. K-N, Amblyrhynchus humerus. L, vector map of all anisotropy. M, vector map of anisotropy of 0.65 and above. N, vector map of the mid region of the bone in the coronal plane with anisotropy 0.65 or higher. O, Anisotropy frequency histogram of Amblyrhynchus humerus...... 131 Figure 5.6. Anisotropy vector maps and histograms of extant aquatic femora. A-D Cryptobranchus femur. B, vector map of all anisotropy. C, Vector map of anisotropy 0.65 and above. D, Vector map of the middle region in the coronal plane with anisotropy 0.65 and above. E, Anisotropy frequency histogram of Cryptobranchus femur. F-I, Ornithorhynchus femur. G, vector map of all anisotropy. H, Vector map of anisotropy 0.65 and above. I, Vector map of the middle region in the coronal plane with anisotropy 0.65 and above. J, Anisotropy frequency histogram of Ornithorhynchus femur. K-N, Amblyrhynchus femur. L, vector map of all anisotropy. M, vector map of anisotropy of 0.65 and above. N, vector map of the mid region of the bone in the coronal plane with anisotropy 0.65 or higher. O, Anisotropy frequency histogram of Amblyrhynchus femur...... 133 Figure 5.7. Anisotropy vector maps and histogram of a Eusthenopteron humerus. B, vector map of all anisotropy. C, vector map of anisotropy 0.65 and above in the flexor half of the bone. D, vector map of anisotropy 0.65 and above in the extensor half of the bone. E, anisotropy frequency histogram of Eusthenopteron humerus...... 134 Figure 5.8 Anisotropy vector maps and histograms of fossil forelimb elements. A-D, YPM.PU.23545 whatcheerid humerus. B, vector map of all anisotropy. C, Vector map of anisotropy 0.65 and above in the flexor half of the bone. D, Vector map of anisotropy 0.65 and above in the extensor half of the bone. E, Anisotropy frequency histogram of YPM.PU.23545. F-I, NSM.007.GF.004.733 Tulerpeton-like ulna. G, vector map of all anisotropy. H, Vector map of anisotropy 0.65 and above in the flexor half of the bone. I, Vector map of anisotropy 0.65 and above in the extensor half of the bone. J, Anisotropy frequency histogram of NSM.007.GF.004.733. K-N, NSM.014.GF.036.002 Acanthostega-like radius. L, vector map of all anisotropy. M, vector map of anisotropy 0.65 and above in the flexor half of the bone. N, vector map of anisotropy 0.65 and above in the extensor half of the bone. O, Anisotropy frequency histogram of NSM.014.GF.036.002...... 135 Figure 5.9. Anisotropy vector maps and histograms of fossil femora. A-D, NSM.005.GF.045.048-A embolomerous femur. B, vector map of all anisotropy. C, Vector map of anisotropy 0.65 and above in the flexor half of the bone. D, Vector map of anisotropy 0.65 and above in the extensor half of the bone. E, Anisotropy frequency histogram of NSM.005.GF.045.048-A. F-I, NSM.007.GF.004.630 indet femur. G,

xii vector map of all anisotropy. H, Vector map of anisotropy 0.65 and above in the flexor half of the bone. I, Vector map of anisotropy 0.65 and above in the extensor half of the bone. J, Anisotropy frequency histogram of NSM.007.GF.004.630...... 137 Figure 5.10. Anisotropy vector maps and historgrams of partial fossil femora. A-D, NSM.005.GF.045.044 Ossinodus-like distal femur. B, vector map of all anisotropy. C, Vector map of anisotropy 0.65 and above in the flexor half of the bone. D, Vector map of anisotropy 0.65 and above in the extensor half of the bone. E, Anisotropy frequency histogram of NSM.005.GF.045.044. F-I, NSM.005.GF.045.047 whatcheerid distal femur. G, vector map of all anisotropy. H, Vector map of anisotropy 0.65 and above in the flexor half of the bone. I, Vector map of anisotropy 0.65 and above in the extensor half of the bone. J, Anisotropy frequency histogram of NSM.005.GF.045.047. K-N, NSM.005.GF.045.047 whatcheerid proximal femur. L, vector map of all anisotropy. M, vector map of anisotropy 0.65 and above in the flexor half of the bone anisotropy 0.65 and above in the flexor half of the bone. N, vector map of anisotropy 0.65 and above in the extensor half of the bone. O, Anisotropy frequency histogram of NSM.005.GF.045.047...... 139 Figure 5.11. Anisotropy vector maps and histograms of fossil fibulae. A-D, NSM.005.GF.045.112 Crassigyrinus-like fibula. B, vector map of all anisotropy. C, Vector map of anisotropy 0.65 and above in the flexor half of the bone. D, Vector map of anisotropy 0.65 and above in the extensor half of the bone. E, Anisotropy frequency histogram of NSM.005.GF.045.112. F-I, NSM.005.GF.045.061 Pederpes-like fibula. G, vector map of all anisotropy. H, Vector map of anisotropy 0.65 and above in the flexor half of the bone. I, Vector map of anisotropy 0.65 and above in the extensor half of the bone. J, Anisotropy frequency histogram of NSM.005.GF.045.061...... 141 Figure 5.12. Anisotropy vector maps and histograms of fossil tibiae A-D, RM.20.5222 Proterogyrinus- like tibia. B, vector map of all anisotropy. C, Vector map of anisotropy 0.65 and above in the flexor half of the bone. D, Vector map of anisotropy 0.65 and above in the extensor half of the bone. E, Anisotropy frequency histogram of RM.20.5222. F-I, NSM.014.GF.036.004 “type 2” whatcheerid tibia. G, vector map of all anisotropy. H, Vector map of anisotropy 0.65 and above in the flexor half of the bone. I, Vector map of anisotropy 0.65 and above in the extensor half of the bone. J, Anisotropy frequency histogram of NSM.014.GF.036.004...... 142 Figure 5.13. PCA plots of all limb elements used. Brown corresponds to terrestrial animals, green to amphibious, blue to aquatic and purple to fossil tetrapod elements. Summary and loadings in table 1 and 2...... 144 Figure 5.14. PCA plots of femora only. Brown corresponds to terrestrial animals, green to amphibious, blue to aquatic and purple to fossil tetrapod elements. Summary and loadings in table 1 and 2...... 146 Figure 5.15. Platypus femur. A, Muscle map adapted from Gambaryan et al (2002). Only the muscles corresponding to the aligned anisotropy vectors are illustrated here. B, vector map of the high anisotropy vectors (0.65 and above) of the whole femur...... 150

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Table of extant animals Genus Common name Felis House Cat Uromastyx Spiny-Tailed Lizard Eublepharis Leopard Gecko Amblyrhynchus Marine Iguana Ornithorhynchus Platypus Lutra River Otter Australian Snapping Elseya Turtle

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Abbreviations

Institutional

Blue Beach Fossil Museum, Hantsport, Nova Scotia, Canada (BBFM); Musée d’Histoire

Naturelle de Miguasha, Québec, Canada (MHNM); Museum of Vertebrate Zoology, University of California, Berkeley, USA (MVZ); Nova Scotia Museum of Natural History, Halifax, Canada

(NSM); Redpath Museum, McGill University, Montreal, Canada (RM); University of Michigan

Museum of Zoology, Ann Arbor, Michigan, USA (UMMZ); Yale Peabody Museum of Natural

History, New Haven, Connecticut, USA (YPM); Museum für Naturkunde Berlin, Germany

(ZMB).

Anatomical

ab, adductor blade; ac, adductor crest; cn, cnemial crest; cp, central prominence; der, dorsal elongate crest; ff, fibular facet; fo, fibular fossa; ft, fourth trochanter; icf, intercondylar fossa; it, internal trochanter; itf, intertrochanteric fossa; mr, mesial ridge; pa, popliteal area;

PIFE, Puboischiofemoralis externus; PIFI, Puboischiofemoralis internus; tf, tibial facet.

Variables

Ani.M, mode of frequency of anisotropy; BV, bone volume; BV/TV, bone volume fraction; Ct.Ar/Tt.Ar, cortical area fraction; Max, global maximum asymptote; Min, global minimum asymptote; P, global transition; Rmax, angular maximum asymptote; Rmin, angular minimum asymptote; RP, angular transition point; RS, angular 1/slope; point; S, global 1/slope;

TV, total volume.

Chapter 1: Introduction Introduction

The Devonian-Carboniferous transition occurred 360 million years ago. Sarcopterygian

(lobe-finned) fish arose in the Devonian period. This group includes the panderichthyids, a group with proximal bony elements in their fins proposed to be homologous to modern tetrapod elements (Boisvert et al., 2008). By the end of the Devonian the first digited tetrapods

Acanthostega, Ichthyostega, and Tulerpeton were present in aquatic environments. The subsequent animals from the Carboniferous Era ranged from aquatic to terrestrial, but exactly how the transition to land took place remained a mystery because of a prominent gap in the fossil record. This gap is now being populated by fossils, but until recently paleontologists were unable to reliably infer locomotor behaviour of early fossil tetrapods from proxies other than external morphological features (Germain and Laurin, 2005; Kriloff et al., 2008; Laurin et al., 2011;

Sanchez et al., 2012, 2013; Quemeneur et al., 2013; Plasse et al., 2019). Analyses of internal structure of limb bones, such as midshaft analyses, show that the internal structure of limb bones provides a robust way to predict locomotor behaviour of tetrapods. Here I expand on that by using three-dimensional data from the internal structures of tetrapod limb bones from Blue

Beach, Nova Scotia, to infer locomotor potential and behaviour in early tetrapods.

In this thesis a “limb” is an appendage consisting of a stylopod, zeugopod, and most importantly a digited autopod. Locomotor behaviour refers to habitual locomotion in a general

(aquatic, amphibious, or terrestrial) environment, and tetrapod is used to describe any animal with four limbs.

Historic hypotheses

Historically the fin-to-limb and water-to-land transitions were thought to be contemporaneous events. The “drying pools hypothesis” for example, imagined sarcopterygian fish traversing small areas of land in order to get from small pools to bigger pools for supporting life (Inger, 1957; Barrell et al., 2011). The idea was that lobe finned fish with more limb-like fins successfully crossed the terrestrial landscape and were able to reproduce. Under the drying pools hypothesis any animal with digited limbs should be terrestrial, or at least amphibious. The discovery of internal gills in Acanthostega (a digited tetrapod) forced the re-evaluation of this traditional idea (Coates and Clack, 1991).

Romer’s Gap

Research into the fin-to-limb transition is complicated by a paucity of material in the fossil record during the time when it was thought that the transition from water to land took place. Romer’s Gap, a phenomenon named for the man who observed it, Alfred Sherwood

Romer (Romer, 1956; Coates and Clack, 1995), originally ranged from the end of the Devonian to the Late Carboniferous. For Romer, Ichthyostega and Acanthostega represented Devonian aquatic animals from the beginning of the fin-to-limb transition and Seymouria, Archeria, and

Eryops represented later, more amphibious body forms from the Carboniferous to Permian

(Romer, 1956). Romer noted the existence of poorly preserved embolomere material in the

Carboniferous but doubted the accuracy of reconstructions at the time of his publication in 1956.

This left most of the Carboniferous period deficient in fossil material. Mid to late Carboniferous tetrapods slowly filled the latter portion of Romer’s Gap in the second half of the twentieth century (Holmes, 1984; Panchen, 1985; Romer and Godfrey, 1989; Panchen and Smithson,

1990; Lombard and Bolt, 1995); however, known isolated elements remained unstudied (see

remarks on Blue Beach material in Lebedev and Coates 1995; Clack and Carroll 2000; Warren and Turner 2004; Coates et al., 2008) until descriptions (Smithson and Clack, 2013; Anderson et al., 2015) and morphotype association by Anderson et al. (2015).

In the early 2000s Romer’s Gap was rapidly filled on both sides by a diverse assemblage of tetrapod material (Clack, 2001, 2002, 2006; Clack and Ahlberg, 2004; Warren and Turner,

2004; Clack and Finney, 2005; Smithson and Clack, 2013, 2018; Anderson et al., 2015; Clack et al., 2018). These fossils refuted other proposed explanations of Romer’s Gap, including a period of low oxygen (Ward et al., 2006), and an extended post extinction recovery period (Sallan et al.,

2010) as well as additional evidence, such as charcoal from wildfires, which require high levels of oxygen in the atmosphere (Mansky and Lucas, 2013). The currently emerging view is that a collection bias towards complete articulated material was responsible for Romer’s Gap

(Smithson et al., 2012).

Blue Beach

Blue Beach, Nova Scotia, is currently the oldest known locality within Romer’s Gap. It is early-mid to mid Tournaisian in age, extending further back than the CM (claviger - macra) biostratigraphical palynozone of similar localities in Britain (Utting et al., 1989; Martel and

Gibling, 1996). However, the Ballagan Formation of Scotland may include some of the slightly older PC (pretiosus - clavata) palynozone (Waters et al., 2009 ongoing). The Ballagan Formation

(which includes a number of sites; see Clack et al., 2017 supplementary information) yields articulated early tetrapod material from the Tournaisian. The dating of the Scottish formations is currently under review (TW:eed Project) and regions of the Ballagan Formation may actually be older than Blue Beach (Waters et al., 2009; Bennett et al., 2016; Clack et al., 2017).

Blue Beach is located along an estuary off the Bay of Fundy near Windsor, Nova Scotia.

Heavy tidal action at the site frequently exposes new fossil material (Mansky and Lucas, 2013).

At the time of deposition Blue Beach was a marginally marine environment, ranging from onshore aerially exposed, to off-shore facies described as lagoonal (Tibert and Scott, 1999;

Mossman and Grantham, 2008), and often stormy (Mansky and Lucas, 2013). These conditions explain the presence of aquatic, amphibious, and possibly terrestrial vertebrates at the same locality. Animals living (or that died) in a nearshore but terrestrial environment could be washed away from nearby locations and deposited at Blue Beach. Aquatic animals washed ashore in previous storms, which had time to decompose between events could subsequently be deposited at Blue Beach as well. Trackways suggest the presence of terrestrial taxa (Mossman and

Grantham, 2008; Mansky and Lucas, 2013). The high energy depositional setting in conjunction with the disarticulated but minimally worn material suggest the animals decomposed before they were transported and deposited at Blue Beach (Mansky and Lucas, 2013). Limb bones such as humeri and femora are disarticulated but identifiable approximately to ordinal level and by morphotypes described in Anderson et al (2015). These identifications will be applied to the material used in this study.

Blue Beach preserves several elements that are similar to Devonian taxa Acanthostega and Tulerpeton (Anderson et al., 2015), which have been previously described as aquatic based on the presence of internal gills, and the depositional setting of the material (Coates and Clack,

1991; Lebedev and Coates, 1995). Blue Beach also has a diverse array of animals similar to some of the material known from the early Carboniferous, presumed to be more terrestrial. There appears to be a community of whatcheeriid-like animals that includes both aquatic (Whatcheeria;

Lombard and Bolt, 1995) and more terrestrial (Pederpes; Clack and Finney, 2005) tetrapods.

Other Carboniferous morphotypes include elements comparable to more amphibious Eoherpeton

(Smithson, 1985) and Proterogyrinus (Holmes, 1984).

Such a range of morphotypes and expected locomotor behaviours makes Blue Beach an ideal locality for investigations into limb bone structure and morphology associated with the evolution of terrestrial locomotion during the fin-to-limb transition. Additionally, there is a known footprint record at Blue Beach, suggesting the likelihood of finding fossils of at least partially terrestrial animals (Mossman and Grantham 2008; Mansky and Lucas 2013). The potential diversity of terrestrial animals will have implications for interpretations of the sequence of events critical to transition onto land. For example, the Blue Beach animals could provide clarification if a single group of animals transitioned onto land and then radiated, or if a diverse group of animals appear to contemporaneously transition to a terrestrial lifestyle. Additionally, the site controls for time so the animals present at Blue Beach represent diverse individuals coexisting at one time potentially across a range of environments, rather than changing fauna or ancestor-descendant relationships (i.e., they do not represent a single lineage changing over time).

In addition to a range of diverse morphotypes identifiable by external morphology, the

Blue Beach tetrapod elements are excellently preserved in three-dimensions. This is critical for the analyses used in chapters four and five, as internal morphology preserves more robust signals of locomotor behaviour than external morphology in many fossil elements (Zeininger et al.,

2016; Su and Carlson, 2017; Kivell et al., 2018; Dunmore et al., 2019; Georgiou et al., 2019)

Historical proxies for locomotion

To transition from an aquatic locomotor behaviour to a terrestrial one, the orientation of fins and their musculature must change. The fins of aquatic fish-like tetrapods must modify from a lateral paddling movement through a viscous medium, to a ventrally-oriented, crawling movement on land. The change in limb orientation from lateral to ventrolateral should be reflected in a change in direction of muscle forces acting on the limbs. Limbs used for locomotion on land experience the force of gravity in a more unidirectional manner as opposed to hydrodynamic forces acting equally in all directions. Additionally, gravity and ground reaction forces should result in forces concentrated on the proximal and distal ends of limb bones, indicating gravitational forces acting across joints.

The main reason proxies are used to infer physiological or biomechanical information such as locomotor behaviour is that soft tissues involved in such processes are not preserved.

Organic material of the bone breaks down and is replaced with mineral precipitates, except in rare instances (e.g. laggerstatten). The primary historical proxy for locomotor behaviour was the presence of digits. The presence of internal gills in addition to a digited limb in Acanthostega forced the need to investigate other proxies for locomotor behaviour. Subsequently proposed proxies include: digit number, limb proportion, degree of ossification, and proportion of compact bone to trabecular bone (Coates and Clack, 1990; Laurin et al., 2004, 2011; Warren and Turner,

2004; Clack and Finney, 2005; Quemeneur et al., 2013).

In Devonian tetrapods the number of digits on a limb was variable (Coates and Clack,

1990). Devonian stem tetrapods Acanthostega, Ichthyostega, and Tulerpeton, were polydactylous, with more than five (up to eight in Acanthostega; Clack, 2006) digits. Coates and

Clack (1990) concluded that pentadactyly (limbs with five digits) was not the primitive state for

early tetrapods, but it was later in the Carboniferous that pentadactyly became the predominant state for tetrapods (Coates and Clack, 1990). This pattern of dactyly was proposed as a proxy for lifestyle inference (Coates and Clack, 1990). More digits (polydactyly) could be used to infer aquatic animals because increased surface area was better for paddling limbs (Smithson et al.,

2012). However, it is unclear if pentadactylous limbs are a necessary precursor to terrestriality, or if terrestrial tetrapods share the pentadactylous condition because selection acted on it after tetrapods transitioned onto land (i.e., plesiomorphy or parallelism).

Limb bone ratios are another proxy used to indicate potential for terrestrial locomotion

(Clack, 2001; Warren and Turner, 2004; Warren, 2007; Sanchez et al., 2010). Shorter proximal elements are associated with aquatic tetrapods, whereas shorter distal elements relative to the proximal element are associated with terrestriality (Clack, 2001; Warren and Turner, 2004;

Warren, 2007; Sanchez et al., 2010). However, conclusions drawn from limb bone proportions may contradict those drawn from dactyly, robusticity, degree of ossification, and histological analysis (Sanchez et al., 2010).

Another proxy for assessing locomotor capabilities is robusticity (Lombard and Bolt,

1995; Milner and Lindsay, 1998; Clack, 2002; Clack and Finney, 2005; Warren, 2007; Shubin et al., 2014). Robusticity is poorly defined, qualitative, and subject to interpretation. In many cases it can be synonymized with well preserved or thick (Clack, 2001, 2002; Daeschler et al., 2006;

Pierce et al., 2012; Shubin et al., 2014). Robusticity is always used in conjunction with another proxy.

The degree of ossification of a tetrapod limb bone is also used to infer lifestyle (Laurin et al., 2004; Coates et al., 2008; Sanchez et al., 2010). Weakly ossified limb bones are associated with aquatic lifestyles, whereas more heavily ossified bones are associated with terrestrial

locomotion because they can withstand greater strain without breaking (Laurin et al., 2004;

Sanchez et al., 2010). Ontogeny may confound these assessments because juveniles have smaller and incompletely ossified limbs regardless of locomotor behaviour (Long and Gordon, 2004;

Warren, 2007).

In Early Carboniferous tetrapods the ossification of limbs, in conjunction with the limb proportion and thickness of cortical bone, are used for assessment of terrestriality (Long and

Gordon, 2004; Coates et al., 2008; Shubin et al., 2014). However, they do not always lead to the same conclusions about locomotor capability (Warren, 2007).

Observations about ossification and robusticity are qualitative descriptions in the early tetrapod literature and lack rigorous testing to associate them with known biological indicators of locomotor behaviour. Paleohistology and biomechanics are more biologically informed routes by which locomotor behaviour is assessed. A number of recent paleohistological investigations reveal patterns of internal anatomy in the limb bones of extant and extinct tetrapods, including thickness of cortical bone and long bone compactness, associated with various lifestyles (Laurin et al., 2004; Kriloff et al., 2008; Canoville and Laurin, 2009; Sanchez et al., 2010; Laurin et al.,

2011; Quemeneur et al., 2013; Sanchez et al., 2016; Molnar et al., 2017; Sanchez et al., 2017;

Molnar et al., 2018; Plasse et al., 2019). In extant tetrapods thicker layers of cortical bone are associated more with terrestrial animals, whereas a large proportion of trabecular bone is associated with an aquatic lifestyle (Kriloff et al., 2008; Canoville and Laurin, 2009; Laurin et al., 2011; Quemeneur et al., 2013; Plasse et al., 2019).

The biological response of trabecular bone to repetitive force, is to modify the bone structure, through modeling or remodelling, to maximize strength and minimize mass of the bone undergoing the repetitive force (Lieberman, 1997)This modelling and remodelling can be

quantified using the degree of anisotropy. Assessing forces from repetitive behaviour, such as ground reaction forces, on bone is used more often in paleoanthropological literature (Ryan and

Ketcham, 2002a, 2002b, 2005; Zeininger et al., 2016; Su and Carlson, 2017; Williams-Hatala et al., 2018). Similar techniques have not been used for studying the fin-to-limb transition in early tetrapods.

The extant phylogenetic bracket (EPB; Bryant and Russell, 1992; Witmer, 1995) is another method used to infer the presence of soft tissues and, by extension, potential locomotor capabilities. The EPB relies on extant bracket taxa to infer the presence of a soft tissue, with varying degrees of confidence, in a fossil taxon. The bracket taxa should be composed of two extant outgroups and an extant sister group to the fossil taxon of interest (Figure 1.1). The outgroup bracket for early tetrapods is composed of the lungfish, plus that group’s outgroup

(Figure 1.1) the coelacanths. The extant sister group to early tetrapods includes all living amphibians and amniotes, which are a diverse group of animals. Paleontologists use the EPB to examine the location and differentiation of musculature on the bone (Hutchinson and Gatesy,

2000; Hutchinson, 2001a, 2001b, 2002; Carrano and Hutchinson, 2002; Molnar et al., 2017,

2018). For early tetrapods the EPB requires comparisons between the fin of a lungfish (very highly derived among living animals) and coelacanth, and often a salamander or iguana. Using the EPB, locations of muscle attachment to fossil bone are inferred and patterns of muscle differentiation are observed across the fin-to-limb transition (Molnar et al. 2017; Diogo et al.

2016; Molnar et al. 2018). When applied to early tetrapods this method requires the assumptions that 1) phylogenies of early tetrapods are well resolved and, 2) osteological correlates are present and homologous from lungfish to amphibian and amniote taxa being used to bracket. This clearly

Figure 1.1. The outgroups and sister group to early tetrapods that are required to infer soft tissues using the extant phylogenetic bracket. involves multiple levels of assumptions and leads to tenuous conclusions when using the EPB in early tetrapods.

External morphological data from fossil material tells an incomplete story. Internal data that reflect biological responses of bone to forces exerted on them are useful to further support conclusions about locomotor capability. Observations of trends based on known biological and physiological responses of bone to behaviours associated with locomotor behaviour are critical for making new meaningful proxies. Additionally, such proxies need not rely on the EPB, or qualitative descriptions such as robusticity.

The purpose of this study is to use multiple qualitative and quantitative methods to analyse the tetrapod limb bones from Blue Beach. I will attempt to establish which morphotypes

were more aquatic or more terrestrial. I expect the diverse faunal assemblage at Blue Beach to reflect a combination of aquatic, amphibious, and terrestrial morphotypes. The animals at Blue

Beach similar to those suggested to be more amphibious, such as Pederpes (Clack and Finney,

2005), Proterogyrinus (Holmes, 1984) and Eoherpeton (Smithson, 1985), should have internal limb bone features that differentiate them from aquatic morphotypes such as those associated with Acanthostega (Coates, 1996) and Tulerpeton (Lebedev and Coates, 1995). Coming from near the base of Romer’s Gap, the are presumed to be close to the origin of terrestriality. The research impact of this study includes establishing the morphotype identity of new bone elements, and identifying variables signalling locomotor behaviour of animals at Blue Beach.

These variables can then be used to study fossils from elsewhere in order to better constrain statements regarding the evolution of terrestriality.

This study will 1) describe new tetrapod material from Blue Beach; 2) attempt to infer differentiation and direction of muscle insertions and associate them with a locomotor behaviour;

2) use extensive data from previous studies (Laurin et al., 2004, 2011; Kriloff et al., 2008;

Canoville and Laurin, 2009; Quemeneur et al., 2013) to infer lifestyle of Blue Beach animals from midshaft profiles; and 3) propose a new method to assess trabecular anisotropy over small regions of an entire bone and associate whole bone anisotropy frequency with locomotor behaviour.

Predictions

In the first chapter I will describe previously undescribed material from Blue Beach.

Recognition of new (or old) material can extend lineages both geographically and temporally.

However, the material first needs to be recognized and exposed to the paleontological community.

Muscle attachment scarring will be compared in the second chapter between extant amphibians, amniotes, and the Blue Beach material. I predict that if the mechanics of locomotion changed from operating in an aquatic environment to a terrestrial environment then; 1) muscle scarring patterns and orientations should shift from those more similar to extant aquatic taxa to more consistent with extant terrestrial taxa, 2) major bone features (such as trochanters) should shift from being present in locations consistent with extant aquatic animals to locations more consistent with extant terrestrial animals, and 3) Blue Beach limb bones should have muscle scarring comparable to a range of extant aquatic and extant terrestrial taxa.

The third chapter uses a computer program devised by Girindot and Laurin (2003) in which a two-dimensional slice at the midshaft of a long bone is used to create a quantitative profile. The variables from the program are plugged into a formula (see supplementary binary formula by Quemeneur et al. 2013) to assess aquatic, or not aquatic lifestyles of the animals associated with the bone. I predict that: 1) because a range morphotypes associated with amphibious Carboniferous (Whatcheerid, Proterogyrinus, Eoherpeton) and Devonian aquatic

(Acanthostega, Tulerpeton) tetrapods are preserved at Blue Beach the bones will reflect a range of terrestrial, amphibious, and aquatic lifestyles; 2) forms associated with the Carboniferous morphotypes will have terrestrial/amphibious profiles.

The fourth chapter will explore a new method using the bone analysis module of

Dragonfly (ORS Montréal). I will qualitatively assess where the strongest forces are acting on the Blue Beach fossil limb bones and create a vector map of the orientations of trabecular structure within each bone. Then I will use previously described variables such as bone volume to total volume (BV/TV) and anisotropy, in addition to variables unique to Dragonfly (ORS,

Montreal) such as cortical area fraction (CtA.TtA). I propose a new variable for assessing

locomotor behaviour of limb bones: frequency of anisotropy. I expect that more amphibious taxa such as Pederpes-like, Proterogyrinus-like and Eoherpeton-like tetrapods will have high trabecular orientation 1) in regions where musculature exerts force associated with terrestrial locomotion, and 2) across joints where gravitational and ground reactive forces are transferred along the limb. Finally, I expect aquatic taxa to show less or no concentrated regions of trabecular orientation, and if they do, I anticipate them to be along the midshaft and associated with musculature used to move the limbs in a lateral swimming motion.

The results and conclusions from chapters one through four will be summarized and their implications discussed in the fifth and final chapter.

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Chapter 2: Descriptions of new tetrapod limb elements from Blue Beach, Nova Scotia Introduction

In 1841 Sir William Logan first observed trackways in the Horton Bluff formation. Sir

William Dawson wrote about trackways from the site in the later 1880s, but bones of tetrapods were not recovered until Baird and Carroll worked the site in the 1960s (Carroll et al., 1972;

Sues et al., 2013). Most previous discussions of Blue Beach tetrapods simply note that isolated material is preserved (Clack et al., 2000; Hall, 2007). Only recently has such material been described from other early Carboniferous sites (Lombard and Bolt, 1995; Clack, 2001, 2002;

Warren and Turner, 2004; Clack and Finney, 2005; Warren, 2007). The tetrapod material from

Blue Beach consists of disarticulated, isolated elements, mostly from the limbs. One recent effort to describe some of the isolated limb bones approached it from the perspective of morphotypes

(Anderson et al., 2015), because enough articulated skeletal material from the Earliest

Carboniferous (Tournaisian and Visean) has been described to aid in assigning the isolated elements to higher taxonomic identifications of such reference material (i.e., Pederpes, Clack and Finney, 2005; Whatcheeria, Lombard and Bolt, 1995; Ossinodus, Warren and Turner, 2004;

Eucritta, Clack, 2001). Efforts to increase sampling of the Tournaisian are also currently underway in Scotland (Clack et al., 2018; Smithson and Clack, 2018; Otoo et al., 2019).

The material that follows is of undescribed and unfigured tetrapod fossil remains from the base of Romer’s Gap at Blue Beach Nova Scotia.

Abbreviations

Blue Beach Fossil Museum, Hantsport, Nova Scotia, Canada (BBFM); Nova Scotia

Museum of Natural History, Halifax, Canada (NSM); Redpath Museum, McGill University,

Montreal, Canada (RM); Yale Peabody Museum of Natural History, New Haven, Connecticut,

USA, (YPM).

Geological context

Blue Beach represents one of the earliest known tetrapod bearing formations of the

Tournaisian. Based on palynozonation it dates late Tn2 (late middle-Tournaisian) to early Tn3

(early late-Tournaisian) subzones, (Utting et al., 1989). This is contemporaneous to slightly older than the tetrapod bearing CM (claviger–macra) zone in Europe, which dates to the early tn3 subzone (Utting et al., 1989). Scottish sites of this age have recently revealed articulated tetrapod material (Smithson and Clack, 2013, 2018; Kearsey et al., 2016; Clack et al., 2018).

However, current work re-examining the Tournaisian localities in Scotland suggests sites from the Ballagran formation may extend below the CM to the PC (pretiosus-clavata) palynozone, so articulated tetrapod material from these formations may be older (Waters et al., 2009; Clack et al., 2017, 2019; Otoo et al., 2019).

The Blue Beach depositional environment was first interpreted as a non marine high energy environment (Martel and Gibling 1991). Later, a periodic marine influence was suggested

(Martel et al., 1993; Martel and Gibling, 1996). The Blue Beach depositional environment is now known as a marginally marine location with at least one fully marine transgression (Tibert and

Scott, 1999; Mansky and Lucas, 2013). The tetrapod material is disarticulated and has signs of

exterior weathering. This suggests the animals were transported a short distance before deposition and fossilization (Mansky and Lucas, 2013).

Blue Beach is located between Hantsport and Avonport, Nova Scotia, Canada, where record breaking high tides of over 10m constantly erode and expose new material. Much of the material is preserved exceptionally well in three dimensions despite high wave and tidal action both at the time of deposition and collection. This is critical for analyses of external and internal structures. External pits, scars, ridges, and foramina are visible on the bone surface and can aid in identification of the animal, it’s physiology, and lifestyle.

Material and Methods

Many specimens were prepared by Sonja Wood of the BBFM prior to being borrowed.

Any unprepared or partially prepared specimens were fully prepared out of the matrix by KL using Paleotools microjacks 1-3, a Chicago Pneumatics air scribe, and pin vice. The elements were reinforced during preparation with polyvinyl acetate and carbowax. NSM.007.GF.004.733 was prepared by Alan Lindoe (University of Alberta). To assess if flattening was a taphonomic or true biological feature of the element the limb bones were scanned in the Xradia Versa 510 at the University of Calgary with settings of 140kV, 72µA, no filter, and magnification 0.4x.

Results

Humeri

Anderson et al. (2015) previously described the humeri YPM.PU.23545 and

YPM.PU.20754, which were borrowed from the Yale Peabody Museum (YPM; New Haven,

Connecticut).

Ulna

NSM.007.GF.004.733

The element (Figure 2.1) is a right ulna and measures 34.3mm in length. It is similar in size to the ulna of Pederpes (Clack and Finney, 2005) from the tip of the prominent olecranon process to the distal surface. This element is heavily worn, but the internal structure reveals minimal crushing. The proximal region of the bone is dominated by the olecranon process, which extends approximately 8mm beyond the rest of the proximal articulating surface. The lateral and mesial edges are concave. A ridge (mr; Figure 2.1 C) runs along the anterior edge of the flexor surface of the element. No other prominent rugosity or features are distinguishable on the bone surface. The element is teardrop shaped in proximal view (Figure 2.1 E) and slightly U-shaped in distal view (Figure 2.1 F). The element is more gracile than ulnae of Acanthostega and

Ichthyostega, but not as elongate as those of Archeria or Greerepeton (Romer, 1957; Romer and

Godfrey, 1989; Coates, 1996; Jarvik, 1996). The anterior edge of the ulna is more concave than in Tulerpeton (Lebedev and Coates, 1995), and the waist at the midshaft is more distinct. The element may be comparable to the ulna of Crassigyrinus (Panchen, 1985), but it has a narrower midshaft with respect to the proximal and distal ends than the ulna figured in Panchen.

Additionally, the ulna of Crassigyrinus does not have a prominent olecranon process preserved.

The ulna of Pederpes also lacks a prominent olecranon process and is less curved along the anterior edge than the ulna of Tulerpeton or NSM.007.GF.004.733 (Lebedev and Coates, 1995;

Clack and Finney, 2005).

Figure 2.1. Micro computed tomography (µCT) images of NSM.007.GF.004.733: right ulna. Tulerpeton- like ulna in (A) extensor, (B) anterior, (C) flexor, (D) posterior, (E) proximal and (F) distal view. mr, mesial ridge. Scalebar 10mm. The element most closely resembles the ulna of Tulerpeton for the reasons outlined above.

Radii

NSM.014.GF.036.002

The element (Figure 2.2 A-F) is 32.7mm from proximal to distal end. It is elongate and the flexor, extensor, anterior, and posterior edges are minimally convex approaching straight.

The midshaft is nearly square in outline (seen in transaxial microcomputed tomography

(microCT) image slices), becoming cylindrical towards the proximal end. The flexor surface displays the most rugosity (Figure 2.2 C). It has striations oriented proximodistally radiating from the long furrow located near the distal midshaft. Both flexor and extensor surface are slightly concave. The flexor surface is bound by a sharp anterior ventral radial crest (Figure 2.2

C, D) and a sharply angled posterior edge. The edge between the posterior and extensor surfaces has a distal prominence with a convex curve in extensor view. The proximal articulating surface

Figure 2.2. µCT images of NSM.014.GF.036.02, right Acanthostega-like radius (A-F) and NSM.007.GF.004.767, left radius (G-L) in (A, G) extensor, (B, H) posterior, (C, I) flexor, (E, K) proximal and (F, L) distal view. vrc, ventral radial crest. Scalebar 10mm. is a hollow, circular, concave shape and the distal articulating surface is flat and D-shaped.

MicroCT scans reveal the element is excellently preserved in three-dimensions.

Based on the circular proximal articulating surface, thin ventral radial crest, and acutely angled edges along a straight shaft, the element most closely resembles the radius of

Acanthostega.

NSM.007.GF.004.767

This element, like NSM.014.GF.036.002 (Figure 2.2 G-L), has a nearly square profile at the midshaft seen in transaxial microCT slices. It is slightly smaller at 29.0mm. The ventral

radial crest is a thin flat projection where the flexor and anterior surfaces meet (Figure 2.2 G, I,

J). No rugosity is discernable on the ventral surface (Figure 2.2 I). The flexor surface is slightly concave, compared to the convex extensor surface. The proximal articulating surface is more cylindrical than the distal end in (transaxial sections); however, the distal end in flexor view is more rounded. The proximal articulating surface is sub oval (Figure 2.2 K), and the distal articulating surface crescent-shaped (Figure 2.2 L).

The element also appears to most closely resemble the radii of Acanthostega, or possibly the more gracile Archeria (Romer, 1957). Internal analysis shows the element is well preserved in three dimensions. Interestingly, it has a very thick cortical layer of bone, whereas

NSM.007.GF.004.767 does not.

Femora

NSM.005.GF.045.048-A

This is a left femur (Figure 2.3 A-F) that is well preserved in three dimensions. It is

60.6mm long and exhibits heavy wear on the adductor blade. No trochanters are clearly visible as a result of the wear. What remains of the adductor blade is separated from the midshaft by a vertical crack. The specimen appears crushed at the distal end, but the midshaft has excellent three-dimensional preservation revealed in transaxial microCT slices. The intertrochanteric fossa is located proximo-ventrally and is a teardrop shape that is highly rugose. Rugose pits are smaller, and closer to circular at the proximal curved end of the teardrop shape. They are deeper and more elongate at the V-shaped end of the intertrochanteric fossa near the midshaft. Distally on the ventral surface there is a sub-triangular region of rugosity within the popliteal area. The popliteal area has more elongate scarring than that observed within the intertrochanteric fossa.

The dorsal surface is smooth and cylindrical at the midshaft. No prominent rugosity or processes

Figure 2.3. µCT images of NSM.005.GF.045.048-A, left embolomerous femur (A-F), and RM.20.5222, a right Acanthostega-like) femur (G-L) in (A, G) extensor, (B, H) anterior, (C, I) flexor, (D, J) posterior, (E, K) proximal, and (F, L) distal views. ab, adductor blade; ac, adductor crest; icf, intercondylar fossa; itf, intertrochanteric fossa; pa, popliteal area. Scalebar 10mm. are present. The anterior surface has numerous small pits converging at the midshaft. The posterior surface is highly concave compared to the anterior. The proximal articulating surface appears oval, is unfinished (lacks a continuous ossified layer of bone) and reveals the internal trabecular structure. The distal articulating surface is an elongate oval with a distinct ridge on the ventral surface of the midline. The ridge has a corresponding concavity (intercondylar fossa) on the dorsal side of the distal surface. The ridge does not clearly divide the articulating surface. In dorsal view there is no discernable bulbous swelling as in the femur of Tulerpeton.

The element is similar to the description of Femur Type 2 from Anderson et al. (2015), which would suggest an embolomerous animal. The posterior profile in ventral view is extremely

convex with respect to the anterior profile in ventral view. Additionally, the dorsal profile appears flat like Femur Type 2.

RM.20.6711

This femur was borrowed from the Redpath Museum (Montréal) for this study. The ventral surface is figured in Clack and Carroll (2000); additional surfaces are illustrated in Figure

2.3 G-H (this work).

A teardrop-shaped intertrochanteric fossa covers most of the proximal end of the ventral surface. The intertrochanteric fossa has smaller pits near the proximal convex end and large elongate pits near the V-shaped end at the midshaft. The adductor blade is worn, although not to the same degree as in NSM.005.GF.045.048-A. The adductor crest provides a posterior edge to the popliteal area which also contains small circular and deep elongate pits. The dorsal surface is smooth and cylindrical. The dorsal profile in anterior or posterior view is sigmoidal, similar to the femur of Acanthostega (Coates 1996). Unlike in NSM.005.GF.045.048-A this element does not appear flattened at the proximal or distal ends. The anterior surface is less concave compared to the posterior surface, but not to the same extent as in NSM.005.GF.045.048-A. The adductor blade of RM.20.6711 does not extend distally as far as in the femur of Acanthostega. Like

NSM.005.GF.045.048-A there are small pits located all over the anterior surface. The posterior surface is smooth. The proximal articulating surface is oval, and the distal articulating surface

V-shaped.

The element shares many similarities to the femur of Acanthostega but is quite different to femur morphotype 4 described in Anderson et al. (2015). The general shape and size are similar to Blue Beach femur NSM.005.GF.045.048-A, but details such as the sigmoidal dorsal

Figure 2.4. µCT images of NSM.007.GF.004.630 a right Tulerpeton-like femur in (A) extensor, (B) anterior, (C) flexor, (D) posterior, (E) proximal, and (F) distal view. ab, adductor blade; ac, adductor crest; cp, central prominence; ff, fibular facet; fo, fibular fossa; itf, intertrochanteric fossa; pa, popliteal area; tf, tibial facet. Scalebar 10mm. surface, and less convex posterior surface suggest and association of RM.20.6711 with

Acanthostega.

NSM.007.GF.004.630

A left femur 65.5mm long (Figure 2.4), is excellently preserved in three dimensions with negligible crushing. The element has a narrower waist than the previously described femora from this study. The proximal end is bulbous in shape and there is approximately 40 degrees of rotation between the proximal and distal articulating surfaces. A crack bisects the element at the midshaft, but what remains of the adductor blade is attached unlike in NSM.007.GF.004.630.

The dorsal surface appears rugose although it is difficult to tell if it is due to preparation, wear, or is a real biological feature. The proximal half of the dorsal surface is very round and moderately wide. There is a ridge similar to the central prominence (cp) described in the femur of Tulerpeton (Lebedev and Coates, 1995). The element is cylindrical at the midshaft before two distal condyles emerge. The condyles extend an equal distance dorsally. The articulating surface of the anterior condyle projects slightly anteriorly (i.e., not in a horizontal plane, approximately

45 degrees from horizontal) whereas the posterior condyle projects distally (i.e., horizontal). The intercondylar fossa is shallow and broad. It extends less than a third of the way up the bone and ends near the distal articulating surface. The ventral surface is also rugose, but much of it looks to be from wear, as exposed trabecular bone is visible. Most of the cortical bone of the distal and posterior edges of the intertrochanteric fossa is worn, exposing trabecular bone beneath. The anteroproximal boundary has a small border of ossified cortical bone (finished bone; arrow in

Figure 2.4 B). The proximal portion of the intertrochanteric fossa is composed of relatively smooth finished bone. The distal teardrop-shaped end is worn exposing the trabecular bone beneath. Most of the adductor blade is absent. The portion of the blade that remains is a triangular projection extending ventrodistally. Exposed trabecular bone suggests the proximal part of the adductor blade broke off rather than the triangular projection reflecting a biological feature. No trochanters are distinguishable. The adductor crest is composed of finished bone. It runs from the distal edge of the adductor blade towards the distal articulating surface and posterior of the midline. The adductor crest and adductor blade meet at right angles with the blade projecting ventrally. The popliteal area is rugose and posteriorly lined by the adductor crest. Large deep elongate foramina on are located where the popliteal area meets the adductor crest. The finished bone preserved on the anterior surface of the midshaft has numerous small pits (see Figure 3.5). The anterior surface is less concave than the posterior surface. The posterior surface preserves little finished bone with the exception of the midshaft, which preserves a small area covered with finished bone. Striations radiate proximally from the posterior surface of the midshaft, but they are obscured by wear. The proximal articulating surface viewed proximally is teardrop shaped. The surface is composed of exposed trabecular bone. The distal articulating surface when viewed dorsal side facing up is a deltoid shape. The intercondylar fossa extends

Figure 2.5. µCT images of YPM.PU.20103 a right Tulerpeton-like/whatcheeriid femur (A-F) and NSM.005.GF.045.042 a left Tulerpeton-like femur (G-L) in (A,G) extensor, (B,H) anterior, (C,I) flexor, (D,J) posterior, (E,K) proximal, and (F,L) distal views. scalebar 10mm. distally from the dorsal surface providing a small amount of finished bone between the tibial and fibular facets. However, ventrally the facets are connected by exposed trabecular bone.

The narrow cylindrical waist and bulbous proximal end make it unlike any other femur I have observed. It shares a proximodorsal tuberosity with the femur of Tulerpeton, a remarkably thin midshaft with the femur of Ossinodus, and is most like Blue Beach specimen

YPM.PU.23545 (Type 1 femur).

NSM.005.GF.045.042

This is a 68.25mm long left femur (Figure 2.5 G-L, and Figure 2.6 E-H) partially prepared by SW at the BBFM before completed by KL at the University of Calgary. In dorsal view the element looks well-preserved; however, preparation of the ventral surface revealed significant crushing of the specimen. Much of the anatomy, including a well-preserved adductor blade remains. However, the ventral surface is crushed into the marrow cavity and the cortical bone is cracked. Overall the element is minimally waisted with very little expansion or broadening at the proximal and distal ends. Axial twist is small but discernable, similar to the femur of Tulerpeton (Lebedev and Coates, 1995). A slight elongate but small ridge is present along the posterior edge of the proximal half of the dorsal surface (der; dorsal elongate ridge).

The proximal two thirds of the dorsal surface appear free of cracking and preserve irregularly shaped pits across the surface. The distal third exhibits much cracking and crushing but remains unbroken. The intercondylar fossa is prominent, if made deeper by crushing. On the dorsal surface the triangular tip of the fossa extends to the posterior end of the adductor blade on the ventral surface. The fibular facet extends further distally than the tibial. Additionally, the tibial facet projects ventroanteriorly approximately 45 degrees offset from the horizontal, unlike the fibular facet which angles ventroposteriorly. The ventral surface has been crushed but the adductor crest, blade and the ridge creating the posterior edge of the intertrochanteric fossa are composed of finished bone and remain attached to the shaft. The adductor blade is robust, consisting of finished bone. It projects ventrally and remains a subrectangular from proximal to distal ends. The adductor blade runs approximately one third the length of the bone. Remains of the intertrochanteric fossa are distinct and are lined anteriorly and posteriorly with finished bone.

Figure 2.6. µCT images (meshes) of YPM.PU.20103 (A-D) and NSM.005.GF.045.042 (E-H) in (A,E) extensor, (B,F) anterior, (C,G) flexor and, (D,H) posterior views. ab, adductor blade; ac, adductor crest; der, dorsal elongate ridge; ff, fibular facet; fo, fibular fossa; icf, intercondylar fossa; it, internal trochanter; itf, intertrochanteric fossa; pa, popliteal area; tf, tibial facet. Scalebar 10mm. The curved proximal boundary of the tear drop shaped fossa is unfinished. Trabecular structure is visible from the tip of the fossa to the proximal tip of the element. The popliteal area retains rugose large pits. These pits extend into the bone, rather than remain on the bone surface. The

anterior surface is moderately well preserved, but no rugosity or pitting is observed. The posterior surface is cracked, and no rugosity is observed. Both anterior and posterior surfaces are slightly concave. The proximal articulating surface appears to be composed of unfinished bone.

The unfinished bone extends at a shallow angle ventrally towards the proximal end of the intertrochanteric fossa. It is unclear if this is the result of crushing, wear, or a true feature of the bone. The proximal articulating surface is a sub oval D-shape. The distal articulating surface is also composed of exposed trabecular bone that extends at a shallow angle towards the ventral surface. The extension of the distal articulating surface to the ventral surface of the bone is considerably less than that described for the proximal end. The distal articulating surface forms a deltoid shape common to many early tetrapod femora. The anterior edge of the deltoid shape extends further than the posterior edge, which has a comma-like shape. The tibial and fibular facets are not separated by finished bone.

The squared off adductor blade dorsal ridge (perhaps related to the cp), and minimal waisting and flaring make this femur most closely resemble a Type 1 Tulerpeton-like femur.

NSM.005.GF.045.044

This specimen preserves the distal two thirds of a femur (Figure 2.7 A-E) measuring

50.05mm in length. The element appears to be cut diagonally at the distal end of the midshaft; it is cut at such an angle so that most of the distal adductor blade remains. The midshaft is more waisted and the distal end of the bone appears broader than in the femora of both Tulerpeton

(Lebedev and Coates, 1995) and Acanthostega (Coates, 1996). The adductor blade has a bifurcation at the proximal end and several foramina are located within. This bifurcation appears consistent with the distal edge of the intertrochanteric fossa.

What remains of the dorsal surface is moderately eroded. The distal condyles are slightly better preserved. The anterior condyle projects anteriorly whereas the posterior condyle projects distally. The anterior condyle has a sigmoidal curve in dorsal view, a feature not observed in any of the femora described above. No similar curve is noted in the femora of Tulerpeton,

Whatcheeria, or Ossinodus. Compared to NSM.007.GF.004.630 and NSM.005.GF.045.048-A the posterior condyle of NSM.005.GF.045.044 projects further distally in relation to the anterior condyle. It also has a smaller sigmoidal curve in dorsal view. The anterior condyle is much larger and broader than the posterior condyle. The intercondylar fossa projects less proximally compared to the other femora; however, erosion of the cortical layer obscures the proximal tip of the fossa. The intercondylar fossa in NSM.005.GF.045.044 extends ventrally at the distal end of the dorsal surface, which creates a trough when the element is viewed distally. The trough extends approximately halfway across the distal articulating surface. The most proximoventral region of the adductor blade appears to have remains of the intertrochanteric fossa. The distal end of the blade (4th trochanter) appears unfinished; however, the unfinished region is nearly perfectly oval similar to the 4th/internal trochanter of the femur of Pederpes (Clack and Finney,

2005), or the 4th trochanter of the femur of Ossinodus. The meeting of the adductor blade and adductor crest forms a gentle concave curve (seen best in Figure 2.8 D). This is unlike the L- shaped curve where the adductor blade meets the adductor crest described in most of the above femora. The popliteal area is a triangular depression with many foramina (large irregularly shaped pits) perforating the distal portion. Little rugosity appears on the anterior or posterior surfaces. Both surfaces have concave profiles. The fibular fossa is more depressed than in any of the Blue Beach femur foramina considered for this analysis and is a sub rectangular shape with

Figure 2.7. µCT images of NSM.005.GF.045.044, distal end of a left Ossinodus-like femur (A-E), and NSM.005.GF.045.047 proximal (K-O) and distal (F-J) segments in (A,F,K) extensor, (B,G,L) anterior, (C,H,M) flexor, (D,I,N) posterior, (E,J) distal and (O) proximal views. ab, adductor blade; ac, adductor crest; cp, central prominence; ff, fibular facet; fo, fibular fossa; ft, fourth trochanter; itf, intertrochanteric fossa; pa, popliteal area; tf, tibial facet. Scalebar 10mm. perforating foramina throughout the depression. The distal articulating surface is deltoid-shaped, a feature shared by many of the Blue Beach femora, and most early tetrapod femora.

The narrow waist, projecting oval shape of the fourth trochanter, and sharp angle between the fibular and tibial facets indicate that this was from an Ossinodus-like animal.

NSM.005.GF.045.047

This element is broken into two pieces (Figure 2.7 F-O), and a 5-10mm section between the two is missing. The proximal piece is 25.4mm long and the distal piece measures 38.3mm.

The adductor blade is heavily eroded, as is the distal end of the ventral surface. Each “half” is well preserved in three dimensions.

The proximal segment (Figure 2.7 K-O) is composed of a small amount of the proximal tip of the adductor blade, intertrochanteric fossa and proximal articulating surface. The dorsal surface shows signs of heavy wear and exposed trabecular bone. The dorsal surface is rounded with the apex of the curve located posterior to the midline. On the ventral surface the intertrochanteric fossa is surrounded by finished bone on the anterior, distal and posterior edges.

There are small pits and foramina located within. The proximal articulating surface is oval shaped.

The dorsal surface of the distal end (Figure 2.7 F-J) is waisted at the midshaft and distally flares out more than in the femur of Tulerpeton (Lebedev and Coates, 1995). The anterior condyle flares anteriorly 45 degrees from horizontal while the posterior condyle extends distally and faces more posteriorly. This is like the configuration observed in NSM.005.GF.045.044. The intercondylar fossa does not extend on the dorsal surface to a location comparable with the adductor blade on the ventral surface. The fossa is obscured by damage at the posterior edge of the distal end. The most concave portion of the intercondylar fossa extends to the distal articulating surface, partially separating the tibial facet from the fibular facet. The distal end preserves most of the adductor blade and crest. The blade is heavily worn so that is consists of almost entirely unfinished bone. A sharp ridge where the blade meets the adductor crest is composed of cortical bone with no exposed trabecular bone. Distally the crest and popliteal areas are eroded. Striations radiate from the midshaft of the bone to the distal end of the adductor crest, the fibular facet, and anterodistal edge of the bone. The distal articulation surface is deltoid to V-

shaped. This element appears similar to the Ossinodus-like femur NSM.005.GF.045.044 because of the thin waist, and angle between fibular and tibial facets.

Previously described

The femur YPM.PU.23550 is described in Anderson et al. (2015), and femora

YPM.PU.20103 (Figure 2.5 A-F, 6 A-D, this study) and RM.20.6711 (Figure 2.3 G-K, this study) are figured but not described in Clack and Carrol (2000). All three femora were borrowed from their respective institutions for this study. Recent investigations in Scotland have revealed elements very similar to the Blue Beach fauna. YPM.20.20103 is remarkably similar in size and shape to the femur NHMUKR 5601 from Inchkeith (Smithson and Clack, 2013), although the

Inchkeith femur appears to be lacking an adductor blade.

A note on femur YPM.PU.20103

Warren and Turner (2004) observed that the femur of Ossinodus most closely resembles

YPM.PU.20103 and that of Tulerpeton. Anderson et al. (2015) suggested that YPM.PU.23550 is a less crushed morphotype of YPM.PU.20103. However, microCT scans reveal the trabecular structures of both elements are crushed, but the cortical layers are well-preserved. I suggest that both share similarities with the femora of Tulerpeton and Ossinodus/Whatcheeria and that

YPM.PU.23550 remain as a Type 1 tulerpetontid morphotype, while YPM.PU.20103 be assigned a new, more whatcheeriid-like morphotype (Femur Type 5).

Fibulae

NSM.005.GF.045.061

This element clearly resembles the fibula of Pederpes (Clack and Finney, 2005).

NSM.005.GF.045.061 (Figure 2.8 A-F) is 37.6mm long measured between proximal and distal

articulation surfaces. The element is waisted, and the distal end more anterodistally expanded than the proximal end. The fibula appears flattened along the dorsoventral plane; however, microCT scans reveal it is well preserved in three dimensions. The entre posterior edge of the flexor surface has a ridge curving concavely (pr; Figure 2.8 C). The flexor surface has minimal rugosity radiating posterodistally at its distal end. At the proximoanterior end of the ridge there are fewer but wider pits resembling foramina. The extensor surface is heavily worn but retains elongate oval foramina radiating from the mid-line of the bone towards the anterior and distal edges. Like the fibula Pederpes (Clack and Finney, 2005), this element has no torsion. The proximal articulating surface dips down into the flexor surface, while the distal articulating surface has an upward lip near the anterior edge of the flexor surface. The proximal articulating surface is teardrop shaped while the distal articulating surface is a flattened elongate oval. The element is extremely similar to the Pederpes fibula figured and described Clack and Finney

(2005).

NSM.005.GF.045.063

This fibula (Figure 2.8 G-L) is shorter than NSM.005.GF.045.061 (30mm), more flattened, and lacks a posterior ridge on the flexor surface. The element is waisted, and the distal end is more flared than the proximal end. The proximal articulating surface dips distally extending onto the flexor surface as in NSM.005.GF.045.061 and the fibula of Pederpes (Clack

Figure 2.8. µCT images of NSM.005.GF.045.061 left Pederpes-like fibula (A-F) in and left fibula NSM.005.GF.045.063 (G-L) in (A,G) extensor, (B,H) posterior, (C,I) flexor, (D,J) anterior, (E,K) proximal, and (F,L) distal views. Scalebar 10mm. and Finney, 2005). The flexor surface has very large pits that look like several large foramina radiating anteriorly from the midshaft. The extensor surface is most rugose at the anterior and posterior edges. The element has a slight triangular shaped depression in the distoanterior edge of the extensor surface. The anterior edge is more concave than NSM.005.GF.045.061, but this may be the result of flattening. The proximal articulating surface has a tri-lobed shape. The distal articulating surface is elongate, sub rectangular, and has concave flexor and extensor edges. The outline of the element resembles the fibula shown but not described in Lombard and Bolt (1995; plate 1). It is shorter and wider than the fibulae of both Pederpes and Tulerpeton.

NSM.005.GF.045.112

This Crassigyrinus-like fibula is described in Lennie, Mansky, and Anderson (In Press).

It appears most similar to the fibula of the Carboniferous and potentially secondarily aquatic tetrapod Crassigyrinus scoticus (Panchen, 1985; Panchen and Smithson, 1990; Carroll, 1992a;

Herbst and Hutchinson, 2019). NSM.005.GF.045.112 has less axial twist than the fibula of

Crassigyrinus and may represent a morphotype less specialized for aquatic locomotion.

Tibiae

RM.20.5222

This tibia (Figure 2.9 A-F) appears Proterogyrinus/embolomere-like. It is 33.55mm long and well preserved in three-dimensions. The flexor surface of the element is rugose near the centre and the rugosity lessens at the peripheries in the proximal and distal directions. Unlike

NSM.005.GF.045.320 the flexor surface of this element has a Y-shaped crest observed in the tibiae of Tulerpeton, Ossinodus, Proterogyrinus and NSM.014.GF.36.004 (Y; described as a proximoposterior to distoanterior ridge along the flexor surface; (Holmes, 1984; Lebedev and

Coates, 1995; Warren and Turner, 2004; Anderson et al., 2015). The posterior edge is concavely shaped like NSM.014.GF.036.003, NSM.014.GF.036.004 (Anderson et al., 2015), and the tibiae of Ossinodus (Warren and Turner, 2004), Eoherpeton (Smithson, 1985), and Proterogyrinus

(Holmes, 1984) rather than notched as in NSM.005.GF.045.320. The anterior surface forms sharp edges where it meets both flexor and extensor surfaces. It becomes rounded at the distal end, and the whole surface is sigmoidal in anterior view. The flat anterior surface bears a ridge comparable to the cnemial process of NSM.005.GF.045.320 and the tibia of Ossinodus (Warren and Turner, 2004), or the rugose ridge described by Holmes (1984) in the tibia of

Proterogyrinus. In proximal view the element is sub triangular to elongate oval. The distal

Figure 2.9. µCT images of left tibiae RM.20.2555 (A-F) and NSM.005.GF.045.320 (G-L) in (A,G) extensor, (B,H) anterior, (C,I) flexor, (D,J) posterior, (E,K) proximal and (F,L) distal views. Scalebar 10mm articulating surface approaches the L-shape described in Tulerpeton, Acanthostega, and

Ossinodus (Lebedev and Coates, 1995; Coates, 1996; Warren and Turner, 2004).

The element is perhaps most similar to the tibiae of Eoherpeton or Proterogyrinus but it lacks torsion and is less gracile.

NSM.005.GF.045.320

This tibia (Figure 2.9 G-L) measures 32.5mm from proximal to distal articulation surface.

This element is flattened in the flexor/extensor plane. In flexor view the element has rugose ridges extending in a radial fan-like pattern from the middle of the anterior edge. The extensor

surface has a prominent cnemial crest on the anterior edge near the mid-line of the element, similar to the tibiae of Ossinodus (Warren and Turner, 2004) and Proterogyrinus (Holmes,

1984). There are small pits covering the entire extensor surface of the bone with elongated depressions radiating out proximally and distally. The pits are small and deep near the midshaft, becoming shallower and steeply angled near the proximal and distal ends. Viewed anteriorly the anterior surface has a sigmoidal curve between proximal and distal ends (Figure 9 H). There is no rotation between the proximal and distal articulating surfaces. The posterior profile is convex except for a concave nearly 90 degree notch (M-shaped?) at the midshaft (seen in Figure 2.9 G,

I). The notch is much less pronounced than the waisting observed in the tibiae of Pederpes

(Clack and Finney, 2005) or Ossinodus (Warren and Turner, 2004). The proximal articulating surface is an irregular oval shape, as is the distal articulating surface. The flattening of the element makes it resemble a more Proterogyrinus-like tibia, although it lacks the Y-shaped crest observed on the flexor surface of the tibiae of Tulerpeton, Ossinodus and Proterogyrinus

(Holmes, 1984; Lebedev and Coates, 1995; Warren and Turner, 2004). NSM.005.GF.045.320 only slightly resembles tibia type 2 (NSM.014.GF.36.004) (Anderson et al 2015), which is considered Pederpes or Ossinodus-like. The element resembles a form transitional between

Ossinodus and Proterogyrinus, likely an early embolomerous form.

Both tibiae are more concave along the posterior edge than Crassigyrinus.

NSM.014.GF.36.004 tibia type 2 is previously described in Anderson et al. (2015) and was borrowed from the NSM for this study. It is most consistent with a Pederpes-like or

Ossinodus-like whatcheeriid (Anderson et al., 2015).

Table 2.1 summarizes the elements borrowed for this study and their morphotype associations. Expected locomotor behaviour are included in the table, however proxies for

locomotor behaviour are based on literature which relies on morphology from features other than stylopod and zeugopod elements (Holmes, 1984; Carroll, 1992a; Lebedev and Coates, 1995;

Lombard and Bolt, 1995; Warren and Turner, 2004; Clack and Finney, 2005)

Table 2.1. Borrowed Blue Beach elements and their attributed taxonomic morphologies and lifestyles. New morphotypes are in bold.

Specimen Element Type Comparable To Locomotor Behaviour

YPM.PU.23545 Humerus Type 1 Whatcheeriid (Ossinodus- Amphibious to like/Pederpes-like) Terrestrial

YPM.PU.20754 Humerus Type 2 Embolomerous Amphibious to Terrestrial

NSM.007.GF.004.733 Ulna Type 1 Tulerpetontid Aquatic

NSM.014.GF.036.002 Radius Type 1 Acanthostegid Aquatic

NSM.007.GF.004.767 Radius Type 1 Acanthostegid Aquatic

NSM.005.GF.045.048- Femur Type 2 Embolomerous Amphibious to A Terrestrial

RM.20.6711 Femur Type 4 Acanthostegid Aquatic

NSM.007.GF.004.630 Femur Type 1 Tulerpetontid Aquatic

NSM.005.GF.045.042 Femur Type 1 Tulerpetontid Aquatic

NSM.005.GF.045.044 Distal Type 5 Whatcheeriid (Ossinodus-like) Amphibious to Femur Terrestrial

NSM.005.GF.045.047 Femur Type 5 Whatcheeriid (Ossinodus-like) Amphibious to Terrestrial

YPM.PU.23550 Femur Type 1 Tulerpetontid Aquatic

YPM.PU.20103 Femur Type 5 Whatcheeriid (revised) Aquatic to Amphibious

NSM.005.GF.045.061 Fibula Type 1 Whatcheeriid (Pederpes-like) Amphibious to Terrestrial

NSM.005.GF.045.063 Fibula Type 1 Whatcheeriid (Whatcheeria- Aquatic like)

NSM.005.GF.045.112 Fibula Type 2 Crassigyrinid Aquatic to Terrestrial?

RM.20.5222 Tibia Type 3 Embolomerous Amphibious to Terrestrial

NSM.005.GF.045.320 Tibia Type 3 Embolomerous Amphibious to Terrestrial

Discussion

Initial sampling and descriptions of some of the material at Blue Beach in 2015 exposed a diverse fauna present in the early Carboniferous. The site includes elements similar to multiple

Carboniferous animals such as the embolomerous Proterogyrinus-like and Eoherpeton-like; whatcheeriid Pederpes-like, Ossinodus-like and Whatcheeria-like; and Crassigyrinus-like

(Mansky and Lucas, 2013; Anderson et al., 2015; Lennie et al. In Press). The animals at Blue

Beach are comparable to these taxa which are previously known from disparate temporal ranges

(North America, Britain, and Australia) and cross over the end Devonian mass extinction. Their presence in the Early Carboniferous complicates ideas about a post extinction period of low diversity or faunal turnover between Devonian and Carboniferous periods (Sallan et al., 2010;

Smithson et al., 2012). Whereas some elements are most comparable to a single known taxon

(ex., Tulerpeton-like femur NSM.005.GF.045.042, or Pederpes-like fibula

NSM.005.GF.045.061), others appear to have a suite of features comparable to multiple known

Devonian and Carboniferous tetrapods (ex., YPM.PU.20103, which is comparable to both the

Devonian Tulerpeton (Anderson et al., 2015) and Ossinodus (Warren and Turner, 2004)).

Temporally disparate body forms present at Blue Beach also capture a diverse range of proposed body forms and locomotor behaviour. For example, the tetrapod Crassigyrinus scoticus, known from the Visean of Scotland has been suggested as a secondarily aquatic animal (Carroll, 1992b;

Herbst and Hutchinson, 2019) and Crassigyrinus-like elements (Mansky and Lucas, 2013;

Lennie et al. In Press) at Blue Beach could represent a better terrestrially adapted animal.

Animals comparable to Eoherpeton and Proterogyrinus could represent more terrestrial animals, as both of those taxa have cranial morphology associated with more terrestrial rather than aquatic forms (Holmes, 1984; Smithson, 1985). Additionally, elements similar to those of the early tetrapod Pederpes (which exhibits asymmetry in its manus and was proposed by Clack and

Finney, 2005 as a terrestrially adapted animal), add to the diversity of earliest known potentially terrestrial animals at Blue Beach.

Most of the fossils at Blue Beach preserve rugosity and foramina despite exposure to weathering. This may be informative regarding muscle insertion and inferences about other soft tissue interactions, including blood vessel and nerve paths through or along the bones. It is difficult to determine the depth of the pits/foramina in an external analysis, but this topic will be discussed in later chapters. Small pits appear common on anterior edges of femora, while larger, irregularly shaped pits are present within the intertrochanteric fossae, popliteal areas and fibular fossae. The distal elements also have many rugosities. Large elongate pits were preserved in all the distal elements excluding NSM.007.GF.004.733 and NSM.007.GF.004.767. Prominent ridges and/or crests were also observed in all the distal elements. This likely indicates extensive soft tissue interaction with the distal elements. Prominent adductor blades and crests on the femora likely indicate musculature exerting forces along the shafts of the proximal bones. This is unusual compared to modern amphibians and amniotes where most of the musculature interacts with the proximal and distal ends of the femora (Petermann and Sander, 2013; Molnar et al.,

2018).

The three-dimensional preservation of most of the material will enable study of internal structures of these elements. Improvement of modern methods regarding non-destructive

sampling and increasing crossover between bone biomechanics and the study of fossil material increases the information we can get from these isolated elements. Despite being isolated and worn, these elements may better reveal how animals were locomoting across the fin-to-limb transition, and when terrestrial locomotion evolved.

Conclusion

The diverse collection of tetrapod material present at Blue Beach ranges from Devonian

Acanthostega-like morphotypes to Carboniferous embolomere and anthracosaur-like forms. The elements described here support previous identifications (Anderson et al., 2015) and contribute descriptions of new material consistent with Devonian Acanthostega-like and Tulerpeton-like elements. A Pederpes-like fibula, and recognition of more Ossinodus-like limb morphotypes supports the presence of multiple Carboniferous whatcheeriid forms, likely with more amphibious habits (Clack and Finney, 2005) and coexisting with Devonian-like animals. The presence of a Crassigyrinus-like fibula is especially intriguing because of previous assertions that Crassigyrinus scoticus is a secondarily aquatic animal (Carroll, 1992; Herbst and

Hutchinson, 2019). The Crassigyrinus-like fibula from Blue Beach may represent the intermediate terrestrial form and contributes to the expansion of the known temporal and geographical range of this group of animals. The Blue Beach limb bones have a range of features comparable to animals from disparate temporal and geographical locations. The morphological features on a single Blue Beach limb bone which are consistent with multiple better known taxa may be indicative of transitional morphologies. Alternatively these may represent a last common ancestor of deep diverging relationships between better known taxa. As we gain comparative material, increased sampling of Tournaisian localities will aid in identification of the isolated

Blue Beach elements. This locality’s diverse overlap in Devonian-like and Carboniferous-like

fauna suggest that there was no clear extinction and replacement or faunal turnover between

Devonian and Carboniferous. The range of morphologies of limb elements from the Early

Carboniferous at Blue Beach supports the occurrence of an early diversification of body plans during the fin-to-limb and water-to-land transitions.

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